The goal of Mission Bio, Inc. (formerly Torrent Bio) is to develop instrumentation that will accelerate the investigation of cellular heterogeneity relevan to human health by rapidly and accurately performing single-cell nucleic acid analysis. In Phase I, we developed a new sorting technology, PCR-activated cell sorting (PACS), which is both high in throughput and able to analyze a wide variety of nucleic acid biomarkers with unmatched sensitivity and specificity. The system is in many ways analogous to Fluorescent Activated Cell Sorting (FACS), except that rather than sorting intact cells, PACS isolates single cells into individual aqueous picoliter- volume drops suspended in oil and performs PCR reactions on their lysates. Using multiplexed TaqMan PCR assays, the system is able to interrogate each cell for the expression of specific combinations of transcripts, genomic DNA, mutations or non-coding RNAs, targets not amenable to antibody-based detection. Moreover, Phase 1 results show our system can sort the single cell lysate and allow it to be recovered for additional downstream molecular characterization. The principal advantage of PACS is that it combines the high- throughput single-cell screening and sorting capability of FACS with the sensitivity of PCR. No technology with similar analysis capability or throughput exists in the marketplace. The intellectual merits of the PACS approach stem from the core ability to deliver a rapid and low-cost solution for massively parallel single-cell genetic and transcriptional analysis on large heterogeneous populations of cells. The impact of cellular heterogeneity on biological function and disease is crucially important to questions in human immunology, stem cell biology, infectious disease and cancer research. By analyzing individual cells within a mixed population, it is possible to identify rare cell populations or transient cell states critical to human health nd development that are otherwise unobservable by ensemble measurements. Commercialization of the PACS system will enable a more detailed and comprehensive analysis of the genomes and transcriptomes of these rare cell populations. Our objective in this Phase II proposal is to take the basic Phase I PACS platform and ready it for commercialization by further optimizing sensitivity and workflow efficiency for downstream single-cell sequencing methods. PACS relies on proven molecular biology approaches, advanced microfluidics, and a solid foundation of intellectual property. Phase II funding will result in a needed and technically achievable research and diagnostic tool with high impact potential in the biomedical marketplace.